We are requesting funds to purchase a Nikon A1R laser scanning confocal microscope. We chose this instrument because it specifically suites the needs of our users, many of whom need to image events within live cells. The A1R is equipped with a high-speed resonant scanner that can scan from 30 (at 512x512 pixels) to 420 frames per second (at 512x32 pixels). It also includes 4Ch 2 GaAsP detectors and 2 High Sensitivity low noise PMT's with spectral sensitivity up to 850nm, A1 detection filters 450/50, 535/50, 600/50 + 685/70m, and 4diode lasers that emit at 405nm, 488nm, 561nm, 640nm. The microscope is equipped with a wide range of objectives to meet the imaging needs of several users. This microscope will be used to support the work of Dr. Robert Conhaim who studies the regulation of microvascular perfusion distribution within the lung. He uses confocal microscopy to image the distribution of fluorescent latex particles of specific diameters infused into the pulmonary circulation. These images are used to determine the vessel populations in which pulmonary vasoconstriction occurs. The A1R will also support the work of Dr. Andreas Friedl who studies development of cancer cells in the central nervous system (gliomas), human breast, and lung. He will use the A1R to measure the uptake of the heparan sulfate proteoglycan glypican-1 by glioma cells, and to observe the induction of syndecan-1 (SDC1) expression in stromal fibroblasts of human breast carcinomas. Dr. Friedl will also use the A1R to measure the migration and proliferation of human lung microvascular endothelial cells in a synthetic microenvironmental chamber. Dr. Dawn Davis studies the roles of specific genes in the regulation of growth and survival of pancreatic beta cells that synthesize, store, and release insulin. She will use photo- bleaching techniques and the rapid imaging ability of the A1R to measure cyclin-dependent kinase migration into beta cells. She will also use the A1R to measure fluorescence resonance energy transfer (FRET) of cAMP and other biosensor signaling in those cells. Dr. Vijay Setaluri will use the A1R to investigate the expression of neuronal differentiation markers, and their subcellular localization in melanoma cells. He will also measure the distribution of cell cycle markers and microtubule protein dynamics including mitotic spindle organization in those cells. Dr. Mihaela Teodorescu will use the A1R to measure the distribution of slow- and fast-twitch myosin heavy chain (MHC) isoforms in the tongue muscles of patients who experience obstructive sleep apnea (OSA). She will also determine how inhaled corticosteroids alter the distribution of these isoforms, which affect the strength and fatiguability of tongue during wakefulness and sleep. She will use the A1R to measure the effects of chronic intermittent hypoxia (CIH) on the distribution of collagen isoforms and smooth muscle thickness in the lungs of rats exposed to CIH. CIH is a cardinal feature of OSA. Dr. Michelle Kimple will use the A1R to measure the role of cyclin-dependent kinases (Cdk's) in the secretory pathway of insulin secreting pancreatic beta cells. She will use photo-bleaching techniques, along with the rapid image acquisition ability of the A1R (420 frames/sec), to measure the glucose-dependence of Cdk mobility within specific beta cell regions. She will also use fluorescence resonance energy transfer (FRET) with the A1R to investigate the role of GalphaZ proteins in the inhibition of cAMP signaling in beta cells, and age-associated changes in beta cell function. The long-term objectives of the above projects are to improve veterans' health in a broad range of diseases. These include the effects of acute blood loss and sepsis on the lung (Dr. Conhaim). They also include the effects of cancer on the central nervous system, breast and lung (Dr. Friedl), and in melanoma (Dr. Setaluri). Dr. Teodorescu's project is designed to reduce the morbidity caused by obstructive sleep apena. The goals of the projects of Drs. Davis and Kimple are to better understand the function of pancreatic beta cells in diabetes. All of these projects have significant implications for veterans' health, and the Nikon A1R we are requesting will facilitate the progress of these important projects.